Efficient generation of transgenic mice with intact yeast artificial chromosomes by intracytoplasmic sperm injection
| Autor: | Moreira, Pedro N., Giraldo, Patricia, Cozar, Patricia, Pozueta, Julio, Jiménez, Adela, Montoliu, Lluís, Gutiérrez-Adán, Alfonso |
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| Jazyk: | angličtina |
| Rok vydání: | 2004 |
| Předmět: |
Transgene integrity
urogenital system Sperm-mediated gene transfer Early development Gamete biology Tyrosinase (Tyr) Assisted reproductive technology Artificial chromosomes Genomic locus Gene regulation Intracytoplasmic sperm injection In vitro fertilization Tyrosinase Intracytoplasmic sperm injection (ICSI) reproductive and urinary physiology |
| Zdroj: | Repositorio de Resultados de Investigación del INIA INIA: Repositorio de Resultados de Investigación del INIA Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria INIA Digital.CSIC. Repositorio Institucional del CSIC instname |
| ISSN: | 1528-6029 |
| Popis: | 5 pages, 1 figure.-- PMID: 15286029 [PubMed].-- Printed version published on Dec 2004.-- Open access version available at the publisher's site. The production of animals with large transgenes is an increasingly valuable tool in biotechnology and for genetic studies, including the characterization and manipulation of large genes and polygenic traits. In the present study, we describe an intracytoplasmic sperm injection (ICSI) method for the stable incorporation and phenotypic expression of large yeast artificial chromosomes (YAC) constructs of submegabase and megabase magnitude. By coinjecting spermatozoa and YACs into metaphase II oocytes, we were able to produce founders exhibiting germline transmission of an intact and functional transgene of 250 kilobases, carrying the mouse tyrosinase locus, used here as a reporter gene to rescue the albinism of recipient mice. More than 35% transgenesis was obtained for this YAC transgene. When compared with the pronuclear microinjection standard method, the efficiency of the ICSI-mediated YAC transfer system was significantly greater. In summary, we describe, for the first time, stable incorporation in the host genome and correct phenotypic expression of large DNA constructs mediated by ICSI. Supported by grants OT02-008 and AGL2002-05783 to A.G.A. and Bio2000-1653 and Bio2003-08196 to L.M. from the Spanish Ministry of Science and Technology. |
| Databáze: | OpenAIRE |
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